Investigation of the tonB gene product of Escherichia coli.

The active transport of all ferric chelates and vitamin B12, infection by bacteriophages T1 and ?80, and killing by a group of colicins are dependent upon the tonB gene product. The initial stage in these processes is an energy-independent binding of the 'substrate' to specific receptor proteins in the outer membrane. Subsequent stages of transport require an energized inner membrane and the tonB gene product. The aim of this study was to identify the tonB product in order to investigate some of its properties directly. The use of ?-tonB transducing phages carrying wild type and mutant tonB alleles to program protein synthesis in UV-irradiated cells enabled the tonB product to be identified; a 40 kilodalton protein was no longer synthesised from tonB deletion and insertion mutants. This protein was associated with the sarkosyl soluble, inner membrane fraction of irradiated cells, suggesting that the translocation of ferric chelates etc. from their initial binding sites, at the exterior of the cells, is dependent upon an inner membrane protein. In an attempt to identify the tonB protein in vivo the tonB region was recloned into a runaway replication plasmid vector. Although cloned proteins were identified, the tonB product was not detected using this system. Physical maps were prepared for the phages and plasmids used in this study, and compared with those described by other workers. The approximate position of the tonB gene was also located. The gene products coded by the tonB region were also identified by utilizing an in vitro coupled transcription translation system. Interestingly, the mutant tonB genes programmed the synthesis of truncated polypeptides in this system. Finally, these results and those of other workers are drawn together to present some models for the action of the tonB product.